Three studies are in progress. The purification and characterization of the phosphodiesterase from the interphotoreceptor matrix continues. It has been found that fast, gentle washes of fresh retinas at 0 degrees provide the best starting material with fewest contaminating species. A concanavalin A affinity column has been used as a second step to remove the major contaminant, a glycoprotein. Chromatography on HPLC sizing columns has yielded a final preparation which is approximately 40% pure. Two affinity chromatography steps (protamine agarose and cGMP-sepharose) are being tested to provide the additional purification needed to obtain a homogenous enzyme. An antibody highly specific for the rod outer segment phosphodiesterase cross reacts with the interphotoreceptor matrix phosphodiesterase. The inhibitors bound to each of these two enzymes are interchangeable and inhibit up to 98% of the activity. The enzyme, guanylate cyclase, in the retina is activated by light. The rate and extent of the activation is under study. A microassay has been set up in the "oil well" to measure femtomoles of product. Our initial findings indicate that the enzyme is unstable at room temperature and humidity in freeze-dried sections with a t l/2 of about 12 hours. Fresh sections, properly cared for, will be required for future studies. High energy phosphate compounds are being measured in retinas of dogs bred to develop a retinal dystrophy. Eyes from controls, carriers, and diseased animals of varying chronological age are sectioned, freeze-dried, and the retinas dissected into 8 layers plus tapetum. They are analyzed by micro methods ("oil well" technique). ATP levels drop 5-fold in the tapetum after 5 weeks of age which correlates with the developmental process.